PROJECT SUMMARY: Biospecimen Unit Despite the growing number of screening options, colorectal cancer (CRC) is still the 4th most common incident cancer and the 2nd most common cause of cancer death in the United States. The majority of CRCs develop from colorectal adenomas. Colonoscopy, the most effective prevention strategy, may not prevent carcinoma in as many as 24% of cases. It reduces deaths from CRC in the left colon, but is not proven for right-sided lesions. Although annual recurrence ranges between 19-23%, current surveillance recommendations provide little improvement in identification of individuals at risk. Thus, it is essential to understand the unique molecular phenotypes of high risk adenomas, laying the foundation for better risk stratification, surveillance and precision prevention. The overarching model for understanding CRC progression has been the adenoma-carcinoma sequence. Although this model initially focused on genetic events, it has been expanded to include epigenetic, immunological, and microenvironmental alterations. A comprehensive and spatially defined landscape of these changes during pre-cancerous adenoma progression does not exist. We propose to develop an atlas of the adenoma-carcinoma sequence that expands upon existing models by conducting spatial analysis of the tumor and its ecosystem using single-cell (sc)RNA-seq and multiplex immunofluorescence (MxIF). In addition, we will characterize the newly-defined microbial biofilm across adenoma progression. Because scRNA-seq and biofilm analysis require specialized tissue collection and processing, the appropriate samples do not exist in any other study. Therefore, we will use novel, rigorously tested protocols as the foundation of our innovative Biospecimen Unit. Sessile serrated adenomas/polyps (SSPs) have only been formally recognized as a CRC precursor within the past decade, and they comprise only 6-10% of adenomas. However, they give rise to 10-35% of CRCs. While most studies in the field are based on conventional adenomas, there is an unmet need to improve our understanding of SSP biology and progression, and its comparison to conventional adenomas, which lead to 65-90% of CRC. Developing a comprehensive spatial landscape and further subclassifying adenoma types, for example as biofilm-positive or biofilm-negative, will enhance our understanding of the role of microenvironment, risk stratification, and precision prevention. Thus, the overall goal of this Biospecimen Unit is to establish a biorepository with varied colorectal tissues from normal rectal mucosa from controls to normal mucosa in adenoma patients, from SSP to conventional adenoma at different histopathologic stages, from epithelial cells to surrounding microenvironment biospecimens (i.e. stroma cells and biofilm microbiome), together with rich metadata on both clinical and epidemiologic exposure data for construction of an atlas. We will recruit 1,800 individuals undergoing screening or surveillance colonoscopy and surgical colon resections. We have extensive experience with colonoscopy-based studies, including over 14,000 participants with gastrointestinal tissue and other related biospecimens.